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Clinical Investigations |
Departments of Radiation Oncology [M. M., A. F., W. L., L. M.], Medical Oncology [D. H.], Biostatistics [M. P.], and Experimental Therapeutics [R. H.], Princess Margaret Hospital, Toronto, Canada M5G 2M9; and Departments of Radiation Oncology [M. M., A. F., W. L., L. M.], Medicine [D. H.], and Medical Biophysics [R. H.], University of Toronto, Toronto, Canada
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ABSTRACT |
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INTRODUCTION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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MATERIALS AND METHODS |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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. The maximum tumor size was determined clinically at examination under
anesthesia and ranged from 2–10 cm with a median of 5 cm. Pelvic and
para-aortic lymph node status was evaluated by magnetic resonance
imaging of the pelvis, computed tomographic scan of the abdomen and
pelvis, and/or bipedal lymphangiography and was classified as negative,
positive, or suspicious according to published criteria (10
, 11)
. Pelvic lymph nodes were negative in 61 patients, positive
in 20 patients, and suspicious in 21 patients. Para-aortic nodes were
negative in 91 patients, positive in two patients, suspicious in seven
patients, and not assessable in two patients. All of the patients with
positive or suspicious para-aortic lymph nodes also had positive pelvic
lymph nodes. None of the patients had metastases to other lymph node
groups or distant sites.
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Measurement of IFP and Oxygen Tension.
IFP was measured using a wick-in-needle apparatus, and oxygen tension
was measured using a polarographic needle electrode system
(Eppendorf-Netheler-Hinz, Hamburg, Germany), as described in detail
elsewhere (7, 8, 9)
. All of the measurements were made during
examination under anesthesia with the patients in the lithotomy
position. Anesthesia was administered using i.v. propofol and inhaled
nitrous oxide, and the inspired oxygen concentration was maintained at
40%. Clinical examination and magnetic resonance imaging of the pelvis
were used to assure that the IFP and oxygen measurements were made in
tumor and not inadvertently in adjacent normal tissue.
IFP was measured at one to five spatially separated locations around the circumference of the visible cervical tumor, at a depth of approximately 2 cm from the surface. Four or more measurements were performed in 87 of the 102 tumors. Multiple measurements of IFP are necessary in each tumor to account for regional heterogeneity, as described previously (7) . The mean of the individual IFP measurements was calculated for each tumor and ranged from -2.8 to 48 mm Hg. The mean IFP was <0 mm Hg in only one patient. It is possible that the measurements in this patient were inadvertently made in the normal tissue surrounding the tumor, because negative IFP values are uncommon in tumors (7 , 12, 13, 14, 15, 16, 17, 18) . However, all of the IFP measurements from all of the eligible patients were included in the analysis to avoid potential bias. Tumor IFP was approximately normally distributed among the patients, with mean and median values for the entire cohort of 20 and 19 mm Hg, respectively.
Oxygen tension was measured immediately before IFP at approximately the same circumferential positions in the tumor. Measurements were made along one to seven linear tracks, with 20 to 30 measurements/track. At least four measurement tracks were used in 88% of the tumors. Oxygenation was expressed as the proportion of the individual oxygen measurements along all of the tracks that were <5 mm Hg (hypoxic proportion or HP5). The HP5 in the individual tumors ranged from 0% to 93%, and the mean and median values for the entire cohort were 46% and 47%, respectively.
The IFP and pO2 measurements were well tolerated in all of the patients, and there were no side effects.
Treatment of Cervix Cancer.
Radiation treatment was prescribed according to the policies of the PMH
Gynecological Oncology Group. External beam pelvic radiation was
administered using anterior-posterior fields or a four-field box
technique and 18–25 MV photons. The irradiated pelvic volume
included the primary tumor as well as the internal, external, and lower
common iliac lymph nodes. Typically, a dose of 45 to 50 Gy in 1.8- to
2-Gy daily fractions was prescribed at the isocenter, and a two
half-value-layer posterior attenuator was used to reduce the rectal
dose by 10 to 20%. An external beam pelvic dose of at least 45 Gy was
delivered to 97 of the 102 patients. In addition, external beam
para-aortic radiation was administered concurrently to 20 patients with
involved pelvic or para-aortic lymph nodes at doses ranging from 12.6
to 50 Gy. Chemotherapy was not used.
After external beam radiation, 97 patients were treated with a single intracavitary low-dose-rate or pulsed-dose-rate brachytherapy application using an intrauterine line source alone without colpostats. A dose of 40 Gy at a dose rate of 0.5–0.8 Gy/h was administered 2 cm lateral to the midpoint of the active sources. Brachytherapy was not feasible in five patients because of unfavorable tumor anatomy. These patients received an external beam boost to the primary tumor using a four-field conformal technique to deliver an additional dose of between 10 and 20 Gy.
Patients were followed at three monthly intervals for the first 2 years after completing radiotherapy and at six monthly intervals thereafter. At each visit, the clinical history was updated, and a physical examination, including pelvic examination, was performed. Laboratory and radiographic tests were obtained as required based on clinical findings. The follow-up of surviving patients ranged from 6 months to 5.5 years, and the median follow-up was 2.5 years.
Analysis.
The primary end point of the study was DFS measured from the date of
diagnosis of cervix cancer. Patients who were documented to have
progressive or recurrent disease after treatment or who died of any
cause were counted as events. Patients who failed to respond to
treatment were classified as having a disease-free interval of zero.
Other patients were counted as failures at the time of first
recurrence. Those who remained alive and free of disease were censored
at the time of last follow-up. DFS was estimated using the Kaplan-Meier
method, and survival curves were compared using the log-rank test.
The primary aim of the study was to determine whether IFP measurements
added additional prognostic information to that readily obtained from
routine clinical and radiographic staging investigations. Therefore, a
clinical multivariate prognostic factor model was first developed using
the Cox proportional hazards model with step-wise selection of clinical
covariates and DFS as the end point. IFP was then added to this model
to determine its independent prognostic effect. The relative importance
of IFP and HP5 was examined by repeating the
analysis and adding both IFP and HP5 to the
clinical model. The 
level for rejecting IFP as an important
prognostic factor was set at 0.025 to minimize the risk of a
false-positive result with multiple challenges to the data. Age, size,
pretreatment hemoglobin concentration, IFP, and
HP5 were tested in the Cox model as continuous
variables. Stage, grade, histology, and pelvic lymph node status were
tested as categorical variables. The interaction terms between IFP and
the significant clinical factors were also tested.
Overall survival, defined as the interval from diagnosis to death from any cause, local recurrence, defined as a first recurrence in the irradiated pelvic volume, and distant recurrence, defined as a first recurrence outside of the irradiated pelvic volume, were secondary end points of the study. Local and distant recurrences defined in this way are competing events. Local recurrence cannot by definition occur after distant recurrence and vice versa, although both can occur simultaneously. Kaplan-Meier estimates of the local or distant recurrence-free rate assume true survival data without competing events and, therefore, overestimate the true recurrence rate. To deal with this problem, the cumulative marginal probabilities of local and distant recurrence were calculated using Gray’s method (19) , which accounts for competing risks. Differences between cumulative recurrence curves were tested for statistical significance using the Wald test with robust standard error. Factors predictive of local and distant recurrence were identified using the Cox proportional hazards model adjusted for competing risks as described by Lunn and McNeil (20) . Patients who recurred initially at distant sites were treated palliatively and not subjected to routine pelvic examination or pelvic imaging. Therefore, the predictive value of IFP for secondary pelvic recurrence (after initial distant recurrence) was not evaluated.
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RESULTS |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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, and the relationship between IFP and
HP5 is shown in Fig. 1b
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Univariate analysis identified a significant association between IFP
dichotomized about the median value of 19 mm Hg and the long-term
outcome of patients after radiotherapy. Patients with low and high IFPs
had 3-year DFSs of 68% and 34%, respectively (P = 0.002), as shown in Fig. 2
. DFS was also influenced by clinical factors, including age (
53
versus >53 years; P = 0.0096),
maximum tumor size (5 versus >5 cm;
P = 0.0008), stage (IB/IIA versus
IIB versus IIIB; P = 0.0003),
pelvic lymph node status (negative versus suspicious
versus positive; P < 0.0001),
pretreatment hemoglobin concentration (126 versus >126
g/liter, P = 0.059), and
HP5 (50 versus >50%;
P = 0.0065). The DFS of patients with
suspicious pelvic lymph nodes was not statistically different from that
of patients with negative nodes; therefore, the two groups were
combined (negative/suspicious versus positive) in the
multivariate analyses.
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, and the combined influence of IFP and lymph node status is shown in
Fig. 4
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shows DFS stratified by both IFP and HP5 in
node-negative patients. The multivariate models for DFS are summarized
in Table 2
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, respectively. There was a trend toward
a higher likelihood of local recurrence in patients with IFPs greater
than the median value of 19 mm Hg (38% versus 24% at 3
years; P = 0.068) and a significantly higher
risk of distant recurrence (47% versus 13%;
P = 0.0014). Multivariate analysis showed IFP
to be significantly associated with both local recurrence
(P = 0.011) and distant recurrence
(P = 0.011), independent of tumor size and
lymph node status. The multivariate analyses for local and distant
recurrence are summarized in Table 3
. The fact that IFP appears to be
only marginally predictive of local control by univariate analysis but
strongly predictive by multivariate analysis mainly reflects inherent
differences in the two techniques of analysis. Patients were divided
into two groups above and below the median IFP value for the univariate
analysis as reflected in Fig. 6a
, with loss of some
information about the relationship between IFP and local control.
However, IFP was treated as a continuous variable in the multivariate
analysis with preservation of all of the possible information in
the data set.
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DISCUSSION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The IFP in any tissue is determined by the interaction of a variety of intrinsic physiological parameters that ultimately determine the capillary pressure, the resistance to fluid leakage across the capillary walls, and the resistance to fluid percolation through the interstitium to the periphery of the tumor (1 , 5 , 22 , 23) . IFP is near zero in most normal tissues, regardless of the capillary pressure, because capillary permeability is low. Also, there is an osmotic gradient that favors the flow of interstitial fluid back into the vasculature, and there is a rich lymphatic network that permeates the tissue and drains any residual interstitial fluid into the central venous circulation (24) . In contrast, tumors usually have abnormal, highly permeable capillaries and lymphatic channels that, although present, have impaired function to the point of being unable to adequately drain excess fluid from the interstitium (2 , 3 , 25) . Therefore, fluid that leaks from the vasculature accumulates in the interstitium and distends the elastic interstitial matrix causing the pressure to rise. In many tumors, the interstitial resistance greatly exceeds the capillary wall resistance, leading to a situation where the IFP nearly equals the capillary pressure (26) .
Capillary pressure generally reflects capillary resistance to blood flow, which is higher in tumors than in the corresponding normal tissues for many reasons, including abnormal vessel structure and organization and high capillary blood viscosity (27, 28, 29, 30) . It has been hypothesized that the accumulation of excess fluid in the interstitium of tumors, which is the underlying cause of elevated IFP, may exert forces that lead directly to compression of tumor capillaries and increased flow resistance (4 , 5 , 31) . Another theory suggests that tumor growth in a confined space leads to increased solid tissue pressure, which in turn causes capillary compression, elevated IFP, and reduced tumor blood flow (31, 32, 33) . It is likely that all of these mechanisms contribute to a greater or lesser degree to the high flow resistance in tumors. Differences in IFP from tumor to tumor, by virtue of the fact that IFP closely tracks the average capillary pressure, provide a relative indication of flow resistance and tumor perfusion.
An earlier analysis of the first 77 patients accrued to this study
showed a weak inverse correlation between IFP and tumor oxygenation
(7)
, in keeping with an inverse relationship between IFP
and blood flow. However, with further accrual, the strength of this
association has diminished to the point where it is no longer
significant, as indicated in Fig. 1
. Studies of IFP and oxygenation in
animal tumors have similarly failed to show a correlation between these
parameters (34, 35, 36)
. This suggests that the strong
prognostic effect of IFP seen in this study is not primarily mediated
through differences in tumor oxygenation. However, it does not exclude
the possibility that the high capillary resistance associated with
elevated IFP is one of many factors contributing to the development of
hypoxia in tumors. High IFP is probably indicative of a chronic
limitation in tumor blood flow, superimposed on a background of
multiple other factors that influence oxygenation independent of
perfusion, including hemoglobin oxygen saturation, longitudinal oxygen
gradients within the tumor microvasculature, long oxygen diffusion
distances, and oxygen consumption by tumor cells (37, 38, 39)
.
Random tumor-to-tumor variability in these other parameters would tend
to mask any underlying association between IFP and oxygenation, making
it difficult to detect in clinical or laboratory experiments.
Although it is possible that IFP and Eppendorf measurement provide independent information about different aspects of tumor oxygenation to explain the results observed in this study, it is also possible that IFP provides information about tumor physiology that is largely unrelated to perfusion and pretreatment oxygen status. Many of the characteristics of tumors that lead to elevated IFP, including abnormal vessel structure and organization and high vascular permeability, arise because of unregulated angiogenesis. This suggests that IFP measurements might provide an integrated measure of the physiological consequences of angiogenesis from one tumor to the next and unique functional information that is not provided by other indicators of angiogenesis such as histological vascular density. The prognostic effect of IFP seen in this study may reflect the more aggressive overall behavior of tumors with high angiogenic activity that is mediated through the interaction of a variety of fundamental physiological and molecular mechanisms including impaired tumor perfusion, the development of hypoxia and acidosis, greater genetic instability, and a greater capacity to form distant metastases. In addition, it has been demonstrated recently that vascular endothelial growth factor, a potent angiogenic protein that is important in the development of both the abnormal tumor vasculature and elevated IFP (40) , is induced by ionizing radiation in some tumors and may protect the vasculature from radiation-induced endothelial cell killing under both oxic and hypoxic conditions (40 , 41) . A clearer understanding of the relationship between IFP and the tumor vasculature would help to explain the strong prognostic effect of IFP that was observed in this study.
In summary, our results provide the first evidence that IFP measurements yield important, clinically relevant biological information about cervix cancer that is independent of both standard prognostic factors and tumor oxygenation. Patients in this study were treated with radiotherapy alone, and those with high-IFP tumors were more likely to recur both within the pelvis and at previously untreated distant sites. The results may or may not be applicable to cervix cancer patients treated with radiation and concurrent cisplatin chemotherapy, which is now standard practice in many North American centers, and to other tumors that may differ pathophysiologically from cervix cancer. Additional laboratory and clinical studies are needed to better understand the mechanisms by which IFP predicts survival of patients with cervix cancer who undergo radiotherapy and to evaluate the prognostic effect of IFP measurements in patients treated surgically. These studies are a prerequisite to using IFP measurements as a means of selecting patients for specific treatments aimed at overcoming the adverse prognostic effect of high IFP and improving survival.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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1 Supported by a Program Project Grant from the
National Cancer Institute of Canada with funds from the Terry Fox
Run. 
2 To whom requests for reprints should be
addressed, at Department of Radiation Oncology, Princess Margaret
Hospital, 610 University Avenue, Toronto, Ontario, Canada M5G 2M9.
Phone: 416-946-2124; Fax: 416-946-2111; E-mail: mike.milosevic{at}rmp.uhn.on.ca
3 The abbreviations used are: IFP, interstitial
fluid pressure; PMH, Princess Margaret Hospital; DFS, disease-free
survival; HP5, hypoxic proportion (percentage of oxygen
measurements <5 mm Hg).
4 A. Fyles, M. Milosevic, D. Hedley, M. Pintilie,
W. Levin, L. Manchul, and R. P. Hill. Tumor hypoxia is an independent
predictor of outcome in patients with node negative cervix cancer,
manuscript in preparation.
Received 3/22/01. Accepted 6/28/01.
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-3 fatty acids can improve radioresponse modifying tumor interstitial fluid pressure, blood rheology and membrane peroxidability.. Anticancer Res., 14: 1145-1154, 1994.[Medline]
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